The National Institute of Alcohol Abuse and Alcoholism estimates the prevalence of alcohol abuse at just over 4.6% of the general population (~9.6 million) and of frank alcohol dependency at just over 3.8% of the population (~7.9 million). To date, no single pharmacotherapeutic agent has been found to be effective for all alcoholics. This collaborative venture brings together expertise in computational drug design, structural biology and enzymology of aldehyde dehydrogenase 2, all of which will be focused during the 2 year timeframe of this award on a single goal - discover and develop a highly potent and selective agent with suitable pharmacokinetic properties for the inhibition of aldehyde dehydrogenase 2 for alcohol dependency. Our long term objective is to target ALDH2 with small organic molecules that possess suitable in vivo efficacy and pharmacokinetic profile and could serve as a adjuvant to currently approved pharmacological approaches toward alcohol dependence therapies (eg. naltrexone or acamprosate) in order to improve the clinical outcomes. The fact that not all alcoholics respond positively to a given treatment exemplifies the importance of developing an arsenal of pharmacotherapeutic agents for the treatment of alcohol use disorders and illustrates the importance of identifying new compounds that have the ability to decrease alcohol drinking. It is our hypothesis that a combined treatment regimen with a drug that exerts its effects in a manner orthogonal to either naltrexone or acamprosate will improve treatment outcomes. In our first aim we perform a computational search to identify additional ALDH2 inhibitors. We introduce several improvements to the computational approach including screening significantly larger libraries and targeting alternative sites and conformational states of the enzyme. In the second aim we characterize the activity of these compounds through enzymology and biophysical studies. In our third aim, a series of in vitro assays are performed to assess the pharmacokinetic properties of the most promising compounds that emerge from Aim 2. This work will set the stage for a follow-up study that will consist of a computational combinatorial search in a multidisciplinary effort that will involve chemical synthesis, biochemical and structural evaluation, and animal pharmacokinetic and efficacy studies. The Indiana University is uniquely positioned through a series of Core facilities to facilitate this process. PUBLIC HEALTH RELEVANCE: Mitochondrial aldehyde dehydrogenase (ALDH2) is most commonly associated with its role in alcohol metabolism and catalyzes the NAD+-dependent oxidation of a broad spectrum of endogenous and biogenic aldehydes to their corresponding carboxylic acids. Inhibition of ALDH2 with small molecules that possess suitable pharmacokinetic properties and efficacy could lead to pharmacotherapeutics to assist in the treatment of alcohol dependence.